Structure-Guided Engineering of Amine Dehydrogenase for Chiral Bulky β -Aryl Amine Synthesis: Dynamic Loop-to-Helix Shift and Hydrogen-Bond Network Reconstruction

Abstract Chiral β-aryl amines are critical structural motifs in various bioactive compounds and pharmaceuticals. Amine dehydrogenases (AmDHs) offer a promising sustainable approach to catalyze the asymmetric reductive amination of β-aryl ketones to chiral β-aryl amines. However, the practical application of AmDHs is severely limited by their narrow substrate scope and notably low catalytic efficiency toward multi-substituted β-aryl ketones. Herein, we report an amine dehydrogenase from Aquisalibacillus elongatus (AeAmDH) and its engineered variant M21 (AeAmDHMutant) developed through a structure-guided mutagenesis strategy. Mutant M21 exhibits a significantly expanded substrate scope, efficiently converting multi-substituted aryl, heteroaryl, and aliphatic ketones, achieving up to a 252-fold enhancement in catalytic efficiency for bulky substrates. Molecular dynamics simulations elucidated a synergistic catalytic mechanism involving dynamic modulation of the enzyme’s active pocket and reorganization of hydrogen-bonding networks. Further process intensification enabled the efficient synthesis of six pharmaceutical intermediates with yields of 62–98% and excellent enantioselectivity (ee>99%).